Balancing Scaffold Degradation and Neo-Tissue Formation in In Situ Tissue Engineered Vascular Grafts.

An essential aspect of cardiovascular in situ tissue engineering (TE) is to ensure balance between scaffold degradation and neo-tissue formation. We evaluated the rate of degradation and neo-tissue formation of three electrospun supramolecular bisurea-based biodegradable scaffolds that differ in their soft-block backbone compositions only. Scaffolds were implanted as interposition grafts in the abdominal aorta in rats, and evaluated at different time points (t = 1, 6, 12, 24, and 40 weeks) on function, tissue formation, strength, and scaffold degradation. The fully carbonate-based biomaterial showed minor degradation after 40 weeks in vivo, whereas the other two ester-containing biomaterials showed (near) complete degradation within 6-12 weeks. Local dilatation was only observed in these faster degrading scaffolds. All materials showed to some extent mineralization, at early as well as late time points. Histological evaluation showed equal and non-native-like neo-tissue formation after total degradation. The fully carbonate-based scaffolds lagged in neo-tissue formation, presumably as its degradation was (far from) complete at 40 weeks. A significant difference in vessel wall contrast enhancement was observed by magnetic resonance imaging between grafts with total compared with minimal-degraded scaffolds.

Evaluation of pliable bioresorbable, elastomeric aortic valve prostheses in sheep during 12 months post implantation.

Pliable microfibrous, bioresorbable elastomeric heart valve prostheses are investigated in search of sustainable heart valve replacement. These cell-free implants recruit cells and trigger tissue formation on the valves in situ. Our aim is to investigate the behaviour of these heart valve prostheses when exposed to the high-pressure circulation. We conducted a 12-month follow-up study in sheep to evaluate the in vivo functionality and neo-tissue formation of these valves in the aortic position. All valves remained free from endocarditis, thrombotic complications and macroscopic calcifications. Cell colonisation in the leaflets was mainly restricted to the hinge area, while resorption of synthetic fibers was limited. Most valves were pliable and structurally intact (10/15), however, other valves (5/15) showed cusp thickening, retraction or holes in the leaflets. Further research is needed to assess whether in-situ heart valve tissue engineering in the aortic position is possible or whether non-resorbable synthetic pliable prostheses are preferred.

Ultrastructural Characteristics of Myocardial Reperfusion Injury and Effect of Selective Intracoronary Hypothermia: An Observational Study in Isolated Beating Porcine Hearts.

In acute myocardial infarction (AMI), myocardial reperfusion injury may undo part of the recovery after revascularization of the occluded coronary artery. Selective intracoronary hypothermia is a novel method aimed at reducing myocardial reperfusion injury, but its presumed protective effects in AMI still await further elucidation. This proof-of-concept study assesses the potential protective effects of selective intracoronary hypothermia in an ex-vivo, isolated beating heart model of AMI. In four isolated Langendorff perfused beating pig hearts, an anterior wall myocardial infarction was created by inflating a balloon in the mid segment of the left anterior descending (LAD) artery. After one hour, two hearts were treated with selective intracoronary hypothermia followed by normal reperfusion (cooled hearts). In the other two hearts, the balloon was deflated after one hour, allowing normal reperfusion (control hearts). Biopsies for histologic and electron microscopic evaluation were taken from the myocardium at risk at different time points: before occlusion (t = BO); 5 minutes before reperfusion (t = BR); and 10 minutes after reperfusion (t = AR). Electron microscopic analysis was performed to evaluate the condition of the mitochondria. Histological analyses included evaluation of sarcomeric collapse and intramyocardial hematoma. Electron microscopic analysis revealed intact mitochondria in the hypothermia treated hearts compared to the control hearts where mitochondria were more frequently damaged. No differences in the prespecified histological parameters were observed between cooled and control hearts at t = AR. In the isolated beating porcine heart model of AMI, reperfusion was associated with additional myocardial injury beyond ischemic injury. Selective intracoronary hypothermia preserved mitochondrial integrity compared to nontreated controls.

Failure of decellularized porcine small intestinal submucosa as a heart valved conduit.

OBJECTIVE: Decellularized extracellular matrix made from porcine small intestinal submucosa, commercially available as CorMatrix (CorMatrix Cardiovascular, Inc, Roswell, Ga) is used off-label to reconstruct heart valves. Recently, surgeons experienced failures and words of caution were raised. The aim of this study was to evaluate decellularized porcine small intestinal submucosa as right-sided heart valved conduit in a xenogeneic animal model. METHODS: A pulmonary valve replacement was performed with custom-made valved conduits in 10 lambs and 10 sheep (1 month [3 lambs and 3 sheep], 3 months [3 lambs and 3 sheep], 6 months [4 lambs and 4 sheep]). Valve function was assessed after implantation and before the animal was put to death. Explanted conduits were inspected macroscopically and analyzed using immunohistochemistry and scanning electron microscopy. They also underwent mechanical testing and testing for biochemical composition. RESULTS: All valved conduits were successfully implanted. Five sheep and 2 lambs died due to congestive heart failure within 2 months after surgery. In the animals that died, the valve leaflets were thickened with signs of inflammation (endocarditis in 4). Five sheep and 8 lambs (1 month: 6 out of 6 animals, 3 months: 4 out of 6 animals, 6 months: 3 out of 8 animals) survived planned follow-up. At the time they were put to death, 5 lambs had significant pulmonary stenosis and 1 sheep showed severe regurgitation. A well-functioning valve was seen in 4 sheep and 3 lambs for up to 3 months. These leaflets showed limited signs of remodeling. CONCLUSIONS: Fifty percent of sheep and 20% of lambs died due to valve failure before the planned follow-up period was complete. A well-functioning valve was seen in 35% of animals, albeit with limited signs of tissue remodeling at ≤3 months after implantation. Further analysis is needed to understand the disturbing dichotomous outcome before clinical application can be advised.

Fibrotic aortic valve disease after radiotherapy: an immunohistochemical study in breast cancer and lymphoma patients.

BACKGROUND: Radiation-associated aortic valve (AV) stenosis is frequently seen as a late sequela after thoracic radiotherapy (RT). Although the clinical relationship between thoracic radiotherapy and valvular dysfunction has been established, the process leading to accelerated aortic valve stenosis remains unclear. The aim of this study was to determine whether increased inflammatory cell infiltration, fibrosis, and calcification is present in aortic valves after radiotherapy at the time of aortic valve replacement. METHODS: Stenotic aortic valve specimens from 43 patients were obtained after surgical aortic valve replacement. A total 28 patients had previously undergone radiotherapy for breast cancer or malignant lymphoma. A total 15 patients were included as control. The valve leaflets were assessed by (immuno)histochemistry for inflammatory cell composition (CD3, CD20, CD68, and CD163) and extracellular matrix changes (collagen and calcification). RESULTS: Aortic valve cell density after radiotherapy for lymphoma was markedly decreased when compared with other groups. Irradiated aortic valve show similar (low) degrees of late T and B lymphocyte infiltration as control valves, whereas macrophage marker CD68 was decreased after radiotherapy for breast cancer. Collagen content was increased following radiotherapy. Aortic valves of patients with lymphoma contained significantly less calcified tissue when compared with the other groups. CONCLUSION: High-dose radiation at a young age (patients with lymphoma) results in cell loss and premature fibrotic aortic valve stenosis as opposed to the degenerative calcific stenosis observed in patients with breast cancer. Our findings suggest a possible dose-dependent effect of radiotherapy on aortic valve fibrosis. The active presence of inflammatory cells may be limited to the acute phase after radiotherapy.

In Situ Remodeling Overrules Bioinspired Scaffold Architecture of Supramolecular Elastomeric Tissue-Engineered Heart Valves.

In situ tissue engineering that uses resorbable synthetic heart valve scaffolds is an affordable and practical approach for heart valve replacement; therefore, it is attractive for clinical use. This study showed no consistent collagen organization in the predefined direction of electrospun scaffolds made from a resorbable supramolecular elastomer with random or circumferentially aligned fibers, after 12 months of implantation in sheep. These unexpected findings and the observed intervalvular variability highlight the need for a mechanistic understanding of the long-term in situ remodeling processes in large animal models to improve predictability of outcome toward robust and safe clinical application.

Intraleaflet hemorrhages are a common finding in symptomatic aortic and mitral valves.

INTRODUCTION: Intraleaflet hemorrhage (ILH) has been reported to occur in calcified degenerated aortic valves. At present, no such information is available for mitral valves or for other types of valvular disease. We examined the prevalence, age, and potential source of ILH in a consecutive series of surgically removed aortic and mitral valves, and related the findings to specific types of heart valve pathology. METHODS: A total of 105 aortic (n=85) and mitral (n=20) valves were retrieved from 100 symptomatic patients. Pathological diagnosis was made on photographic images and histology. Presence, extent, and age of ILH; its possible association with calcification; microvessels; and microvascular leakage were assessed with conventional and immunohistochemical staining methods and related to the type of underlying valvular disease. RESULTS: Pathological diagnosis revealed degenerative aortic valve disease (n=70), postinflammatory disease (n=16), endocarditis (n=12), myxoid degenerative mitral valve disease (n=6), and one normal valve. ILH was found in 86% of aortic and 75% of mitral valves. Microvessels were present in 91% of all valves. Microvascular leakage was noted in 70% of aortic and 84% of mitral valves; in both groups, colocalization with ILH was found in 48%. Most aortic valves (91%) contained calcium deposits, of which 54% showed colocalization with ILH. In 66% of valves with ILH, a combination of recent hemorrhage and iron deposits was seen, indicating an ongoing process of episodic hemorrhages. CONCLUSION: The prevalence of ILH is very high in resected heart valves. Both aortic and mitral valves showed an association of ILH with microvessels, microvascular leakage, and calcifications. We speculate that repetitive microvascular-leakage-related ILH may contribute to valve dysfunction on the (very) long term.

Incidental necropsy finding of a quadricuspid aortic valve.

Quadricuspid aortic valve is a rare congenital cardiac malformation often associated with abnormal valve function. In this article, we present a case of quadricuspid aortic valve only diagnosed at the time of post-mortem examination.

In Situ Tissue Engineering of Functional Small-Diameter Blood Vessels by Host Circulating Cells Only.

Inflammation is a natural phase of the wound healing response, which can be harnessed for the in situ tissue engineering of small-diameter blood vessels using instructive, bioresorbable synthetic grafts. This process is dependent on colonization of the graft by host circulating cells and subsequent matrix formation. Typically, vascular regeneration in small animals is governed by transanastomotic cell ingrowth. However, this process is very rare in humans and hence less relevant for clinical translation. Therefore, a novel rat model was developed, in which cell ingrowth from the adjacent tissue is inhibited using Gore-Tex sheathing. Using this model, our aim here was to prove that functional blood vessels can be formed in situ through the host inflammatory response, specifically by blood-borne cells. The model was validated by implanting sex-mismatched aortic segments on either anastomoses of an electrospun poly(ɛ-caprolactone) (PCL) graft, filled with fibrin gel, into the rat abdominal aorta. Fluorescent in situ hybridization analysis revealed that after 1 and 3 months in vivo, over 90% of infiltrating cells originated from the bloodstream, confirming the effective shielding of transanastomotic cell ingrowth. Using the validated model, PCL/fibrin grafts were implanted, either or not loaded with monocyte chemotactic protein-1 (MCP-1), and cell infiltration and tissue development were investigated at various key time points in the healing cascade. A phased healing response was observed, initiated by a rapid influx of inflammatory cells, mediated by the local release of MCP-1. After 3 months in vivo, the grafts consisted of a medial layer with smooth muscle cells in an oriented collagen matrix, an intimal layer with elastin fibers, and confluent endothelium. This study proves the regenerative potential of cells in the circulatory system in the setting of in situ vascular tissue engineering.